Messenger RNAs (mRNAs) are formed in the nuclei of eukaryotic cells by extensive posttranscriptional processing of primary RNA polymerase II transcripts. These transcripts, called heterogeneous nuclear RNAs (hnRNAs) or pre-mRNAs, are associated with specific proteins throughout the time they are in the nucleus. The collective term for the proteins that bind hnRNAs and their nuclear processing products, and which are not stable components of other classes of RNP complexes such as snRNPs, is hnRNP proteins. The hnRNP proteins are avid RNA-binding proteins and the processing of hnRNA in the nucleus takes place in hnRNA-protein (hnRNP) complexes. The significance of hnRNP proteins is that they influence the structure and interactions of hnRNAs and their processing into mRNAs. As several hnRNP proteins shuttle between the nucleus and the cytoplasm, they may also have novel functions in the cytoplasm as well as in nucleo- cytoplasmic transport of mRNAs. The major hnRNP proteins are among the most abundant proteins in the nucleus and hnRNP complexes are thus also of interest because they are major nuclear structures. In order to understand how the post-transcriptional pathway of gene expression operates in the cell, we investigate the structure, function, and localization of the hnRNP proteins. Though much progress has been made in these areas over the past several years, a great deal of fundamental importance remains to be learned. The focus of the research proposed in this grant application is on the further molecular characterization of the hnRNP proteins, including determination of their sequences, structures, RNA-binding activities, protein-protein interactions, assembly into complexes, biochemical activities, and functions in mRNA biogenesis.